Despite advancements in antiretroviral therapy (ART) that reduces the viral load to undetectable levels and improve CD4 T cell counts, viral eradication has not been achieved due to HIV-1 persistence in resting CD4+ T-cells. We, therefore, characterized the tat gene, which is essential for HIV-1 replication and pathogenesis, from 20 virologically controlled aging individuals with HIV (HIV+) on long-term ART and improved CD4+ T-cell counts, with a particular focus on older individuals. Peripheral blood mononuclear cell genomic DNA from HIV+ were used to amplify tat gene by polymerase chain reaction followed by nucleotide sequencing and analysis. Phylogenetic analysis showed that each HIV+tat sequences were confined to their own subtrees and well discriminated from other HIV+ sequences. Moreover, there was a low degree of viral heterogeneity and lower estimates of genetic diversity within these individuals’ tat sequences, which decreased with increasing CD4 T counts in these HIV+. Most HIV+ Tat deduced amino acid sequences showed intact open reading frames and maintained the important functional domains for Tat functions, including transactivation, TAR binding, and nuclear localization. Furthermore, Tat-deduced amino acid sequences showed variation in previously characterized cytotoxic T lymphocytes (CTL) epitopes, suggesting escape mutants. In conclusion, a low degree of genetic variability and conservation of functional domains and variations in CTL epitopes were the features of tat sequences that may be contributing to viral persistence in these 20 aging individuals with HIV on long-term ART.
Get full access to this article
View all access options for this article.
References
1.
HIV Surveillance Report, 2019. Centers for Disease Control and Prevention; 2021.
2.
MayMT, GompelsM, DelpechV, et al.UK Collaborative HIV Cohort (UK CHIC) Study. Impact on life expectancy of HIV-1 positive individuals of CD4+ cell count and viral load response to antiretroviral therapy. Aids, 2014; 28(8):1193–1202; doi: 10.1097/QAD.0000000000000243
3.
MarcusJL, ChaoCR, LeydenWA, et al.Narrowing the gap in life expectancy between HIV-Infected and HIV-Uninfected individuals with access to care. J Acquir Immune Defic Syndr, 2016; 73(1):39–46; doi: 10.1097/QAI.0000000000001014
4.
BehrensNE, WertheimerA, KlotzSA, et al.Reduction in terminally differentiated T cells in virologically controlled HIV-infected aging patients on long-term antiretroviral therapy. PLoS One, 2018; 13(6):e0199101; doi: 10.1371/journal.pone.0199101
5.
BehrensNE, WertheimerA, LoveMB, et al.Evaluation of HIV-specific T-cell responses in HIV-infected older patients with controlled viremia on long-term antiretroviral therapy. PLoS One, 2020; 15(9):e0236320; doi: 10.1371/journal.pone.0236320
6.
RobertsJD, BebenekK, KunkelTA. The accuracy of reverse transcriptase from HIV-1. Science, 1988; 242(4882):1171–1173; doi: 10.1126/science.2460925
7.
DengK, PerteaM, RongvauxA, et al.Broad CTL response is required to clear latent HIV-1 due to dominance of escape mutations. Nature, 2015; 517(7534):381–385; doi: 10.1038/nature14053
8.
ChunTW, EngelD, BerreyMM, et al.Early establishment of a pool of latently infected, resting CD4(+) T cells during primary HIV-1 infection. Proc Natl Acad Sci U S A, 1998; 95(15):8869–8873; doi: 10.1073/pnas.95.15.8869
9.
ChunTW, CarruthL, FinziD, et al.Quantification of latent tissue reservoirs and total body viral load in HIV-1 infection. Nature, 1997; 387(6629):183–188; doi: 10.1038/387183a0
10.
FinziD, BlanksonJ, SilicianoJD, et al.Latent infection of CD4+ T cells provides a mechanism for lifelong persistence of HIV-1, even in patients on effective combination therapy. Nat Med, 1999; 5(5):512–517; doi: 10.1038/8394
11.
ClarkE, NavaB, CaputiM. Tat is a multifunctional viral protein that modulates cellular gene expression and functions. Oncotarget, 2017; 8(16):27569–27581; doi: 10.18632/oncotarget.15174
12.
BerkhoutB, SilvermanRH, JeangKT. Tat trans-activates the human immunodeficiency virus through a nascent RNA target. Cell, 1989; 59(2):273–282; doi: 10.1016/0092-8674(89)90289-4
13.
FeinbergMB, BaltimoreD, FrankelAD. The role of Tat in the human immunodeficiency virus life cycle indicates a primary effect on transcriptional elongation. Proc Natl Acad Sci U S A, 1991; 88(9):4045–4049; doi: 10.1073/pnas.88.9.4045
14.
FaustTB, BinningJM, GrossJD, et al.Making sense of multifunctional proteins: Human immunodeficiency virus type 1 accessory and regulatory proteins and connections to transcription. Annu Rev Virol, 2017; 4(1):241–260; doi: 10.1146/annurev-virology-101416-041654
15.
IzmailovaE, BertleyFM, HuangQ, et al.HIV-1 Tat reprograms immature dendritic cells to express chemoattractants for activated T cells and macrophages. Nat Med, 2003; 9(2):191–197; doi: 10.1038/nm822
16.
SecchieroP, ZellaD, CapitaniS, et al.Extracellular HIV-1 tat protein up-regulates the expression of surface CXC-chemokine receptor 4 in resting CD4+ T cells. J Immunol, 1999; 162(4):2427–2431.
17.
EnsoliB, BuonaguroL, BarillariG, et al.Release, uptake, and effects of extracellular human immunodeficiency virus type 1 Tat protein on cell growth and viral transactivation. J Virol, 1993; 67(1):277–287; doi: 10.1128/jvi.67.1.277-287.1993
18.
CarvalloL, LopezL, FajardoJE, et al.HIV-Tat regulates macrophage gene expression in the context of neuroAIDS. PLoS One, 2017; 12(6):e0179882; doi: 10.1371/journal.pone.0179882
19.
JiangY, ChaiL, FasaeMB, et al.The role of HIV Tat protein in HIV-related cardiovascular diseases. J Transl Med, 2018; 16(1):121; doi: 10.1186/s12967-018-1500-0
20.
MediouniS, DarqueA, BaillatG, et al.Antiretroviral therapy does not block the secretion of the human immunodeficiency virus tat protein. Infect Disord Drug Targets, 2012; 12(1):81–86; doi: 10.2174/187152612798994939
21.
GaneshanS, DickoverRE, KorberBT, et al.Human immunodeficiency virus type 1 genetic evolution in children with different rates of development of disease. J Virol, 1997; 71(1):663–677; doi: 10.1128/jvi.71.1.663-677.1997
22.
LinchangcoGVJr, FoleyB, LeitnerT. Updated HIV-1 consensus sequences change but stay within similar distance from worldwide samples. Front Microbiol, 2021; 12:828765; doi: 10.3389/fmicb.2021.828765
23.
ThomasH, JukesCRC. Mammilian Protein Metabolism. Academic Press; 1969.
24.
WattersonGA. On the number of segregating sites in genetical models without recombination. Theor Popul Biol, 1975; 7(2):256–276; doi: 10.1016/0040-5809(75)90020-9
25.
NeiM. Molecular Evolutionary Genetics. Columbia University Press: New York: 1987.
26.
NeiM, GojoboriT. Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions. Mol Biol Evol, 1986; 3(5):418–426; doi: 10.1093/oxfordjournals.molbev.a040410
27.
RayneF, DebaisieuxS, YezidH, et al.Phosphatidylinositol-(4,5)-bisphosphate enables efficient secretion of HIV-1 Tat by infected T-cells. Embo J, 2010; 29(8):1348–1362; doi: 10.1038/emboj.2010.32
28.
HolmesEC, deAZPM. Genetic drift of human immunodeficiency virus type 1? J Virol, 1998; 72(1):886–887.
29.
TetoG, FonsahJY, TagnyCT, et al.Molecular and genetic characterization of HIV-1 Tat Exon-1 gene from cameroon shows conserved Tat HLA-Binding epitopes: Functional implications. Viruses, 2016; 8(7); doi: 10.3390/v8070196
30.
KuppuswamyM, SubramanianT, SrinivasanA, et al.Multiple functional domains of Tat, the trans-activator of HIV-1, defined by mutational analysis. Nucleic Acids Res, 1989; 17(9):3551–3561; doi: 10.1093/nar/17.9.3551
31.
GarciaJA, HarrichD, PearsonL, et al.Functional domains required for tat-induced transcriptional activation of the HIV-1 long terminal repeat. EMBO J, 1988; 7(10):3143–3147; doi: 10.1002/j.1460-2075.1988.tb03181.x
32.
HauberJ, MalimMH, CullenBR. Mutational analysis of the conserved basic domain of human immunodeficiency virus tat protein. J Virol, 1989; 63(3):1181–1187; doi: 10.1128/JVI.63.3.1181-1187.1989
33.
RappaportJ, LeeSJ, KhaliliK, et al.The acidic amino-terminal region of the HIV-1 Tat protein constitutes an essential activating domain. New Biol, 1989; 1(1):101–110.
34.
WeiP, GarberME, FangSM, et al.A novel CDK9-associated C-type cyclin interacts directly with HIV-1 Tat and mediates its high-affinity, loop-specific binding to TAR RNA. Cell, 1998; 92(4):451–462; doi: 10.1016/s0092-8674(00)80939-3
35.
PierleoniR, MenottaM, AntonelliA, et al.Effect of the redox state on HIV-1 tat protein multimerization and cell internalization and trafficking. Mol Cell Biochem, 2010; 345(1–2):105–118; doi: 10.1007/s11010-010-0564-9
36.
RangaU, ShankarappaR, SiddappaNB, et al.Tat protein of human immunodeficiency virus type 1 subtype C strains is a defective chemokine. J Virol, 2004; 78(5):2586–2590; doi: 10.1128/jvi.78.5.2586-2590.2004
37.
GarberME, WeiP, KewalRamaniVN, et al.The interaction between HIV-1 Tat and human cyclin T1 requires zinc and a critical cysteine residue that is not conserved in the murine CycT1 protein. Genes Dev, 1998; 12(22):3512–3527; doi: 10.1101/gad.12.22.3512
38.
OrsiniMJ, DebouckCM. Inhibition of human immunodeficiency virus type 1 and type 2 Tat function by transdominant Tat protein localized to both the nucleus and cytoplasm. J Virol, 1996; 70(11):8055–8063; doi: 10.1128/jvi.70.11.8055-8063.1996
39.
RuizAP, AjasinDO, RamasamyS, et al.A naturally occurring polymorphism in the HIV-1 Tat basic domain inhibits uptake by bystander cells and leads to reduced neuroinflammation. Sci Rep, 2019; 9(1):3308; doi: 10.1038/s41598-019-39531-5
40.
HeM, ZhangL, WangX, et al.Systematic analysis of the functions of lysine acetylation in the regulation of tat activity. PLoS One, 2013; 8(6):e67186; doi: 10.1371/journal.pone.0067186
41.
Endo-MunozL, WarbyT, HarrichD, et al.Phosphorylation of HIV Tat by PKR increases interaction with TAR RNA and enhances transcription. Virol J, 2005; 2:17; doi: 10.1186/1743-422x-2-17
42.
AddoMM, AltfeldM, RosenbergES, et al.HIV Controller Study Collaboration. The HIV-1 regulatory proteins Tat and Rev are frequently targeted by cytotoxic T lymphocytes derived from HIV-1-infected individuals. Proc Natl Acad Sci U S A, 2001; 98(4):1781–1786; doi: 10.1073/pnas.98.4.1781
43.
YusimK, KesmirC, GaschenB, et al.Clustering patterns of cytotoxic T-lymphocyte epitopes in human immunodeficiency virus type 1 (HIV-1) proteins reveal imprints of immune evasion on HIV-1 global variation. J Virol, 2002; 76(17):8757–8768; doi: 10.1128/jvi.76.17.8757-8768.2002
44.
ChakrabortyS, RahmanT, ChakravortyR. Characterization of the protective HIV-1 CTL epitopes and the corresponding HLA Class I alleles: A step towards designing CTL based HIV-1 vaccine. Adv Virol, 2014; 2014:321974; doi: 10.1155/2014/321974
45.
JonesNA, WeiX, FlowerDR, et al.Determinants of human immunodeficiency virus type 1 escape from the primary CD8+ cytotoxic T lymphocyte response. J Exp Med, 2004; 200(10):1243–1256; doi: 10.1084/jem.20040511
46.
HusainM, HahnT, YedavalliVR, et al.Characterization of HIV type 1 tat sequences associated with perinatal transmission. AIDS Res Hum Retroviruses, 2001; 17(8):765–773; doi: 10.1089/088922201750237040
47.
RonsardL, LataS, SinghJ, et al.Molecular and genetic characterization of natural HIV-1 Tat Exon-1 variants from North India and their functional implications. PLoS One, 2014; 9(1):e85452; doi: 10.1371/journal.pone.0085452
48.
SpectorC, MeleAR, WigdahlB, et al.Genetic variation and function of the HIV-1 Tat protein. Med Microbiol Immunol, 2019; 208(2):131–169; doi: 10.1007/s00430-019-00583-z
49.
AsamitsuK, FujinagaK, OkamotoT. HIV Tat/P-TEFb Interaction: A potential target for novel Anti-HIV therapies. Molecules, 2018; 23(4); doi: 10.3390/molecules23040933
50.
SelbyMJ, PeterlinBM. Trans-activation by HIV-1 Tat via a heterologous RNA binding protein. Cell, 1990; 62(4):769–776; doi: 10.1016/0092-8674(90)90121-t
51.
BehrensNE, LoveM, BandlamuriM, et al.Characterization of HIV-1 Envelope V3 region sequences from virologically controlled HIV-Infected older patients on long term antiretroviral therapy. AIDS Res Hum Retroviruses, 2021; 37(3):233–245; doi: 10.1089/AID.2020.0139
52.
LoveM, SamoraL, BarkerD, et al.Genetic analysis of HIV-1 vpr Sequences from HIV-Infected older patients on long-term antiretroviral therapy. Curr HIV Res, 2022; 20(4):309–320; doi: 10.2174/1570162X20666220705124341
53.
McBrienJB, KumarNA, SilvestriG. Mechanisms of CD8(+) T cell-mediated suppression of HIV/SIV replication. Eur J Immunol, 2018; 48(6):898–914; doi: 10.1002/eji.201747172
54.
DampierW, NonnemacherMR, MellJ, et al.HIV-1 Genetic variation resulting in the development of new quasispecies continues to be encountered in the peripheral blood of well-suppressed patients. PLoS One, 2016; 11(5):e0155382; doi: 10.1371/journal.pone.0155382
55.
KearneyMF, SpindlerJ, ShaoW, et al.Lack of detectable HIV-1 molecular evolution during suppressive antiretroviral therapy. PLoS Pathog, 2014; 10(3):e1004010; doi: 10.1371/journal.ppat.1004010
56.
BozziG, SimonettiFR, WattersSA, et al.No evidence of ongoing HIV replication or compartmentalization in tissues during combination antiretroviral therapy: Implications for HIV eradication. Sci Adv, 2019; 5(9):eaav2045; doi: 10.1126/sciadv.aav2045
57.
WarrenJA, CluttonG, GoonetillekeN. Harnessing CD8(+) T Cells Under HIV antiretroviral therapy. Front Immunol, 2019; 10:291; doi: 10.3389/fimmu.2019.00291
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
